Abstract
Production of reactive oxygen species (hydroxyl radicals, superoxide radicals and hydrogen peroxide) was studied using EPR spin-trapping techniques and specific dyes in isolated plasma membranes from the growing and the non-growing zones of hypocotyls and roots of etiolated soybean seedlings as well as coleoptiles and roots of etiolated maize seedlings. NAD(P)H mediated the production of superoxide in all plasma membrane samples. Hydroxyl radicals were only produced by the membranes of the hypocotyl growing zone when a Fenton catalyst (FeEDTA) was present. By contrast, in membranes from other parts of the seedlings a low rate of spontaneous hydroxyl radical formation was observed due to the presence of small amounts of tightly bound peroxidase. It is concluded that apoplastic hydroxyl radical generation depends fully, or for the most part, on peroxidase localized in the cell wall. In soybean plasma membranes from the growing zone of the hypocotyl pharmacological tests showed that the superoxide production could potentially be attributed to the action of at least two enzymes, an NADPH oxidase and, in the presence of menadione, a quinone reductase.
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Abbreviations
- DEPMPO:
-
5-Diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide
- DPI:
-
Diphenyleneiodonium
- EPR:
-
Electron paramagnetic resonance
- MD:
-
Menadione
- NBT:
-
Nitro blue tetrazolium chloride
- NQR:
-
Naphthoquinone reductase
- POBN:
-
α-(4-Pyridyl-1-oxide)-N-tert-butylnitrone
- RBOH:
-
Respiratory burst oxidase homologue
- SOD:
-
Superoxide dismutase
- XTT:
-
Na,3′-1-[phenylamino-carbonyl]-3,4-tetrazolium-bis(4-methoxy-6-nitro)benzenesulfonic acid hydrate
References
Able AJ, Guest DI, Sutherland MW (1998) Use of a new tetrazolium-based assay to study the production of superoxide radicals by tobacco cell cultures challenged with avirulent zoospores of Phythophthora parasitica var nicotinae. Plant Physiol 117:491–499
Bérczi A, Møller IM (2000) Redox enzymes in the plant plasma membrane and their possible roles. Plant Cell Environ 23:1287–1302
Bielski BHJ, Shiue GG, Bajuk S (1980) Reduction of nitro blue tetrazolium by CO2 − and O2 − radicals. J Phys Chem 84:830–833
Bolwell G, Bindschedler LV, Blee KA, Butt VS, Davies DR, Gardner SL, Gerrish C, Minibayeva F (2002) The apoplastic oxidative burst in response to biotic stress in plants: a three-component system. J Exp Bot 53:1367–1376
Bradford MM (1976) A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye-binding. Anal Biochem 72:248–254
Brightman AO, Barr R, Crane FL, Morré DJ (1988) Auxin-stimulated NADH oxidase purified from plasma membrane of soybean. Plant Physiol 86:1264–1269
Buckhout TJ, Hrubec TC (1986) Pyridine nucleotide-dependent ferricyanide reduction associated with isolated plasma membranes of maize (Zea mays L.) roots. Protoplasma 135:144–154
Chen S, Schopfer P (1999) Hydroxyl-radical production in physiological reactions. A novel function of peroxidase. Eur J Biochem 260:726–735
Cosio C, Dunand C (2009) Specific functions of individual class III peroxidase genes. J Exp Bot 60:391–408
Cross AR (1987) The inhibitory effects of some iodonium compounds on the superoxide generating system of neutrophils and their failure to inhibit diaphorase activity. Biochem Pharmacol 36:489–493
DeHahn T, Barr R, Morré DJ (1997) NADH oxidase activity present on both the external and internal surfaces of soybean plasma membranes. Biochim Biophys Acta 1328:99–108
Döring O, Lüthje S, Böttger M (1992) Inhibitors of the plasma membrane redox system of Zea mays L. roots: the vitamin K antagonists dicumarol and warfarin. Biochim Biophys Acta 1110:235–238
Doussière J, Vignais PV (1992) Diphenylene iodonium as an inhibitor of the NADPH oxidase complex of bovine neutrophils. Factors controlling the inhibitory potency of diphenylene iodonium in a cell-free system of oxidase activation. Eur J Biochem 208:61–71
Finkelstein E, Rosen GM, Rauckman EJ (1982) Production of hydroxyl radicals by decomposition of superoxide spin-trapped adducts. Mol Pharm 21:262–265
Halliwell B, Gutteridge JMC (2006) Free radicals in biology and medicine, 4th edn. Clarendon Press, Oxford
Heyno E, Klose C, Krieger-Liszkay A (2008) Origin of cadmium-induced reactive oxygen species production: mitochondrial electron transfer versus plasma membrane NADPH oxidase. New Phytol 179:687–699
Hruz T, Laule O, Szabo G, Wessendorp F, Bleuler S, Oertle L, Widmayer P, Gruissem W, Zimmermann P (2008) Genevestigator V3: a reference expression database for the meta-analysis of transcriptomes. Adv Bioinform 420747
Janzen EG, Wang YY, Shetty RV (1978) Spin trapping with α-pyridyl 1-oxide N-tert-butyl nitrones in aqueous solutions. J Am Chem Soc 100:2923–2925
Keller T, Damude HG, Werner D, Doerner P, Dixon RA, Lamb C (1998) A plant homolog of the neutrophil NADPH oxidase gp91phox subunit gene encodes a plasma membrane protein with Ca2+ binding motifs. Plant Cell 10:255–266
Kobayashi M, Kawakita K, Maeshima M, Doke N, Yoshioka H (2006) Subcellular localization of Strboh proteins and NADPH-dependent O2–generating activity in potato tuber tissues. J Exp Bot 57:1373–1379
Kuchitsu K, Kosaka H, Shiga T, Shibuya N (1995) EPR evidence for generation of hydroxyl radical triggered by N-acetylchitooligosaccharide elicitor and a protein phosphatase inhibitor in suspension-cultured rice cells. Protoplasma 188:138–142
Lin W (1982) Responses of corn root protoplasts to exogenous reduced nicotinamide adenine dinucleotide: oxygen consumption, ion uptake, and membrane potential. Proc Natl Acad Sci USA 79:3773–3776
Lind C, Cadenas E, Hochstein P, Ernster L (1990) DT-diaphorase: purification, properties and function. Methods Enzymol 186:287–301
Liszkay A, Kenk B, Schopfer P (2003) Evidence for the involvement of cell wall peroxidase in the generation of hydroxyl radicals mediating extension growth. Planta 217:658–667
Liszkay A, van der Zalm E, Schopfer P (2004) Production of reactive oxygen intermediates (O ·−2 , H2O2 and OH·) by maize roots and their role in wall loosening and elongation growth. Plant Physiol 136:3114–3123
Lüthje S, van Gestelen P, Córdoba-Pedregosa MC, Gonzalez-Reyes JA, Asard H, Villalba JM, Böttger M (1998) Quinones in plant plasma membranes—a missing link? Protoplasma 205:43–51
Mika A, Lüthje S (2003) Properties of guaiacol peroxidase activities isolated from corn root plasma membranes. Plant Physiol 132:1489–1498
Mika A, Minibayeva F, Beckett RP, Lüthje S (2004) Possible functions of extracellular peroxidases in stress-induced generation and detoxification of active oxygen species. Phytochem Rev 3:173–193
Mika A, Buck F, Lüthje S (2008) Membrane-bound class III peroxidases: identification, biochemical properties and sequence analysis of isoenzymes purified from maize (Zea mays L.) roots. J Proteomics 71:412–424
Mika A, Boenisch M, Hopff D, Lüthje S (2010) Membrane-bound guaiacol peroxidases from maize (Zea mays L.) roots are regulated by methyl jasmonate, salicylic acid, and pathogen elicitors. J Exp Bot 61:831–841
Minibayeva F, Kolesnikov O, Chasov A, Beckett RP, Lüthje S, Vylegzhanina N, Buck F, Böttger M (2009) Wound-induced apoplastic peroxidase activities: their roles in the production and detoxification of reactive oxygen species. Plant Cell Environ 32:497–508
Mojović M, Vuletić M, Bačić GG, Vučinić Ž (2004) Oxygen radicals produced by plant plasma membranes: an EPR spin-trap study. J Exp Bot 55:2523–2531
Ramos CL, Pou S, Britigan BE, Cohen MS, Rosen GM (1992) Spin trapping evidence for myeloperoxidase-dependent hydroxyl radical formation by human neutrophils and monocytes. J Biol Chem 267:8307–8312
Renew S, Heyno E, Schopfer P, Liszkay A (2005) Sensitive detection and localization of hydroxyl radical production in cucumber roots and Arabidopsis seedlings by spin trapping electron paramagnetic resonance spectroscopy. Plant J 44:342–347
Sagi M, Fluhr R (2001) Superoxide production by plant homologues of the gp91phox NADPH oxidase: modulation of activity by calcium and by tobacco mosaic virus infection. Plant Physiol 126:1281–1290
Sagi M, Fluhr R (2006) Production of reactive oxygen species by NADPH oxidases. Plant Physiol 141:336–340
Schopfer P (2001) Hydroxyl radical-induced cell-wall loosening in vitro and in vivo: Implications for the control of elongation growth. Plant J 28:679–688
Schopfer P, Liszkay A (2006) Plasma membrane-generated reactive oxygen intermediates and their role in cell growth of plants. BioFactors 28:73–81
Schopfer P, Liszkay A, Bechthold M, Frahry G, Wagner A (2002) Evidence that hydroxyl radicals mediate auxin-induced extension growth. Planta 214:821–828
Schopfer P, Heyno E, Drepper F, Krieger-Liszkay A (2008) Naphthoquinone-dependent generation of superoxide radicals by quinone reductase isolated from the plasma membrane of soybean. Plant Physiol 147:864–878
Schweikert C, Liszkay A, Schopfer P (2000) Scission of polysaccharides by peroxidase-generated hydroxyl radicals. Phytochemistry 53:565–570
Schweikert C, Liszkay A, Schopfer P (2002) Polysaccharide degradation by Fenton reaction- or peroxidase-generated hydroxyl radicals in isolated plant cell walls. Phytochemistry 61:31–35
Siegel D, Gustafson D, Dehn DL, Han JY, Boonchoong P, Berliner LJ, Ross D (2004) NAD(P)H:quinone oxidoreductase 1: role as a superoxide scavenger. Mol Pharmacol 65:1238–1247
Simon-Plas F, Elmayan T, Blein J-P (2002) The plasma membrane oxidase NtrbohD is responsible for AOS production in elicited tobacco cells. Plant J 31:137–147
Tarsio JF, Shapiro BL (1984) Competitive inhibition of human mitochondrial NADH dehydrogenase by Cibacron Blue F3GA. Enzyme 32:188–192
Thein M, Michalke W (1988) Bisulfite interacts with binding sites of the auxin-transport inhibitor N-1-naphthylphthalamic acid. Planta 176:343–350
Torres MA, Jones JDG, Dangl JL (2006) Reactive oxygen species signaling in response to pathogens. Plant Physiol 141:373–378
Zancani M, Nag G, Vianello A, Macri F (1995) Copper-inhibited NADH-dependent peroxidase activity of purified soya bean plasma membranes. Phytochemistry 40:367–371
Zhou M, Diwu Z, Panchuk-Voloshina N, Haugland RP (1997) A stable non-fluorescent derivative of resorufin for the fluorimetric determination of trace hydrogen peroxide: applications in detecting the activity of phagocyte NADPH oxidase and other oxidases. Anal Biochem 253:162–168
Acknowledgments
We are grateful to Drs. F. Sparla and P. Trost (Università di Bologna, Italy) for stimulating discussions and provision of the NQR antibodies. Research costs were funded by CEA and CNRS.
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Heyno, E., Mary, V., Schopfer, P. et al. Oxygen activation at the plasma membrane: relation between superoxide and hydroxyl radical production by isolated membranes. Planta 234, 35–45 (2011). https://doi.org/10.1007/s00425-011-1379-y
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DOI: https://doi.org/10.1007/s00425-011-1379-y